Synthesis and evaluation of anti-microbial of some new Biotin carrying sugar moieties

The current research involves preparing biotin sugar derivatives and evaluating their vital activity against some types of microbial, through a set of interactions, where the biotin was initially converted into an ester derivative according to the Fischer method. Then the ester derivative 2 was treated with aqueous hydrazine to prepare the derivative 3. After that, the derivative 3 was condensed with a number of monosaccharides using the Schiff-Base reaction. The prepared compounds were spectrophotometrically characterized via modern methods. Results of the biological tests showed that the derivatives 4-8 possess antibacterial vital activity as well as anti-fungi.


INTRODUCTION
Biotin is a derivative of Imidazole and it is available in natural foods as an essential ingredient and in some foods as a dietary supplement [1]. Also, a large portion of biotin is manufactured by gut bacteria that human cells, as well as many organisms, cannot produce [2]. In the year 1953, Donald B.Melville [3] was synthesized biotin sulfoxide that showed good biological activity, and as well as the great importance of biotin derivatives in various areas of life led to an interest in the synthesis of more new derivatives for it. Either in the year 2018, Heather Nesbitt was concluded that the biotin derivatives which contain the sugar molecules in their composition were used as medicinal compounds against cancer cells in the pancreas [4]. As sugars contribute to the importance of the drug and its penetration and dissolution inside the body [5]. Biotin derivatives were used as an antiinflammatory for certain types of bacteria and good breast cancer [6,7]. As They can also be used as a new drug for cancer and vaccine therapy, as well as the use of ligands that inhibit the action of TLRS receptors, enzyme inhibiting agents, making them resistant to enzymatic and chemical decomposition sometimes because they do not have oxygenic or glycoside binding [8]. They are many sugars that have been studied with various biological activities, including it is immune system [9], antioxidant [10], anti-inflammatory [11]. In this work, we prepared a series of biotin derivatives combined with a number of monosaccharides and were evaluated for their bioactivity.

EXPERIMENTAL SECTION
Melting point were measured using steuart andare uncorrected .FT-IR Spectra using KBr disk were recorded on shimadzu-FT-IR-8400 S Fourier transform , 13 C-NMR , 1 H-NMR Spectra were recorded on a BrukerAvancelll spectrometer at 400 MHz using DMSO-d 6 with TMS reference university Ghazi Osman Pasha in Turkey.all materials used are pure and processed by companies Fluka,Aldrich,GCC.

Chemical synthesis Synthesis of Ethyl 5-(2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl)pentanoate 2[12]
In round pottom flask100 ml with three neck put on heater with magnetic stirrer ,condenser and thermometer with a balloon filled with inert nitrogen gas ,was taken (1g,4.1 mmol) of biotin1 and dissolved in 20ml of absolute ethanol with the addition 3ml of concentrated sulfuric acid and the 2 reaction mixture was reflux for 13h .The progress of reaction was monitored by TLC in A mixture of (MeOH:CH 2 Cl 2 ) with a ratio of (2:8)(v:v)as a mobile phase,after that the solvent residual was evaporated under the vacuum pressure.It was neutralized by adding drops of 10% Na 2 SO 4 with stirring then poured on ice crush.then the product precipitated and filtered through a Buckner funnel and wash with distilled water ,dried and recrystallized with ethanol to give white precipitate. Yield:75%, m.p=151-153 0 C ,Rf=0.42

Synthesis of 5-(2-oxohexahydro-1H-thieno[3,4-d]imidazol-4-yl)pentanehydrazide3[13]
In round bottom flask 100 ml ,was taken (1.7g ,0.0062 mmol) of derivative2biotin ethyl ester dissolved in 50ml of absolute ethanol with the addition of 6ml of hydrazine hydrate and the mixture was reflux for 14h at a temperature 78 0 C .the reaction was monitored by TLC in a mixture of (CHCl 3 :MeOH)in the ratio(9:1)(v:v) As a mobile phase ,the reaction mixture was left to cool at room temperature.After that the precipitate was filtered and washed with ether.The resulting product was purified as a white solid by recrystallizing it from ethanol. Yield: 51% , m.p =242-243 0 C , Rf=0.2

General Method of thePreparationSugar derivatives of biotin 4-8[10]
In round bottom flask 100 ml ,was taken (0.1163g ,0.45mmol) of derivative 3Biotin hydrazide with (0.5mmol)of monosaccharides dissolved in 20ml of absolute ethanol and the mixture was reflux for(4-6)h .the reaction was monitored by TLC in a mixture of (CHCl 3 :MeOH) at a ratio (8:2)(v:v) As a mobile phase ,after the completion of the reaction ,the reaction mixture was left for coolding at room temperature.After that , the resulting product was collected and recrystallized to obtain the sugars derivatives of biotin.

Biological activities
We are conducting evaluation experiment for antimicrobial activity of the synthesized compounds were investigated in vitro against both Gram positive bacteria (Staphylococcus aureus) and Gram negative bacteria (Escherichia coil) as well as Fungi (Candida albicans ,Aspergillusniger)using the broth dilution method [14] preparation of nutrient broth , subculture , base layer medium,a gar medium was according to the standard procedure.The standard and test compounds were dissolved in DMSO to obtained a concentration of 50 Mg/ml and 100Mg/ml.The samples were incubated at 37 0 C for 24h (bacteria) , at 25 0 C for 72h (fungi), zone of inhibition was measured in mm. Cephalexin and Fluconazole were used as standard antibiotics to compare the results

RESULT AND DISCUSSION Chemistry
Ester derivative of biotin2 were prepared via reaction of biotin1with ethanol in acidic medium. Next, ester derivative 2 was condensed with hydrazine hydrateunder reflux to get the derivative 3. Later ,the derivative 3 was condensed with a number of monosaccharides using the Schiff-Base reaction. The synthesis methods to the novel compounds were shown in the following scheme 1.     (8)